This proposal seeks to achieve a durable cure of HIV through the use of autologous gene-modified cells in a 'Defend and Destroy' approach. Our central premise is that autologous cells can be used to eradicate HIV if they are 1. Genetically rendered HIV-uninfectable, and, 2. Enhanced in their activity by specific measures to destroy viral reservoirs. We have therefore assembled a team of investigators with complementary areas of expertise to execute the following specific aims: Specific aim 1: Utilize novel genetic modification technologies to engineer HIV- resistant immune systems with enhanced ability to destroy viral reservoirs. (Derrick Rossi, PI) This takes advantage of expertise in the Rossi lab with the CRISPR- Cas9 genome editing system and modified-mRNA technology. Specific aim 2: Enhance engraftment of gene modified hematopoietic stem cells with reduced toxicity, niche sparing conditioning of the host to minimize morbidity and costs without compromising targeting of viral reservoirs. (David Scadden, PI) Specific aim 3: Test approaches for viral control and vira reservoir depletion in a HIV-infected and anti-viral treated 'humanized' mouse model. (Todd Allen, PI) Specific aim 4: Pre-clinical development of HIV-resistant hematopoietic stem cells. (CRISPR Therapeutics, Rodger Novak, PI) Each aims maps to a project and the projects are supported by two cores: A. Administration and Biostatistics, and, B. Humanized mouse core generating, treating and analyzing BLT mice infected with HIV and treated with triple drug anti-retroviral therapy. Each of the four Specific Aims described above could be conducted independently as a scientifically significant research project. However, it is the synergy achieved through the coordinated accomplishment of these aims that will enable the testing of a multi-pronged strategy for eradicating HIV infection. Testing of such a complex set of strategies, which together represent the platform for a Defend and Destroy approach to curing HIV infection, would simply not be possible if any one project were conducted out of the context of the other three. Thus, it is only the concerted execution of this package of four Projects, supported by two critical Cores, that will facilitate testing of the proposed, highly novel strateg for curing HIV.